Parathyroid, kidney and other cell types sense extracellular calcium (Ca2+O) through a recently cloned, G-protein-coupled Ca2+0-sensing receptor. Several diseases of Ca2+ homeostasis result from point mutations in the human homolog of this receptor, which are thought to either increase or decrease its activity. The mutations are present throughout the receptor protein and potentially provide a valuable source of information on the impact of changes in its primary structure on its expression and/or biological activity. To date however, very few of these mutations have been studied in detail when expressed in heterologous cell systems, particularly mammalian expression systems. The goals of this proposal are four-fold: (1) to introduce several of these mutations into a construct of the human Ca2+0-sensing receptor suitable for site-directed mutagenesis; (2) to express the mutant constructs in the human embryonic kidney cell line, HEK 293 (3) to determine whether the mutations affect the receptor's ability to reach the cell surface, and, if not, (4) to ascertain if they alter its maximal activity and/or apparent affinity for Ca2+ and/or other agonists. The results of this study may help to elucidate not only the basis for normal Ca2+ homeostasis and inherited abnormalities in it but potentially also other disorders in which there is abnormal Ca2+0-sensing, including primary and uremic secondary hyperparathyroidism.